Molecular characterization and functional analysis of thioredoxin peroxidase-2 from Babesia duncani

Parasitol Int. 2026 Oct:114:103290. doi: 10.1016/j.parint.2026.103290. Epub 2026 Apr 27.

Abstract

Human babesiosis caused by Babesia duncani is an emerging zoonotic disease that poses a growing threat to public health. Since the first reported case in 1991, knowledge of this pathogen has remained limited. Recent completion of the B. duncani genome sequence has accelerated research aimed at elucidating its molecular biology. During its intraerythrocytic life cycle, B. duncani is exposed to oxidative stress generated by redox reactions. In this study, we identified and characterized an antioxidant enzyme from B. duncani-thioredoxin peroxidase-2 (BdTPx-2), a member of the peroxiredoxin family. BdTPx-2 gene was amplified from both cDNA and gDNA of B. duncani, revealing an open reading frame of 792 bp with no introns. The native form, cellular localization, and immunogenicity of BdTPx-2 were determined using polyclonal antibodies raised against recombinant BdTPx-2 (rBdTPx-2) and sera from B. duncani-infected mice. Antioxidant activity was assessed using a mixed-function oxidation (MFO) assay. BdTPx-2 was recognized by sera from mice infected with B. duncani or B. microti, dogs infected with B. gibsoni, and water buffalo infected with B. orientalis, indicating that TPx-2 is conserved and exhibits cross-reactivity among Babesia species. A native protein of approximately 35 kDa was specifically detected in B. duncani lysates. The MFO assay demonstrated that BdTPx-2 inhibits the oxidation of plasmid DNA by a DTT/FeCl₃ mixture, preventing its conversion from the supercoiled to the nicked form. In BALB/c mice immunized with rBdTPx-2 and subsequently challenged with Babesia, parasite burden was significantly reduced, accompanied by elevated levels of the proinflammatory cytokine IFN-γ, suggesting a protective immune response induced by rBdTPx-2. This work characterizes TPx-2 as an antioxidant enzyme in B. duncani and provides insights into a key mechanism by which the parasite defends against oxidative stress during its intraerythrocytic stage.

Keywords: Antioxidant enzyme; Babesia duncani; Peroxiredoxin; Thioredoxin peroxidase-2; Zoonotic disease.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Babesia* / enzymology
  • Babesia* / genetics
  • Babesia* / immunology
  • Babesiosis* / immunology
  • Babesiosis* / parasitology
  • Buffaloes
  • Dogs
  • Female
  • Humans
  • Mice
  • Mice, Inbred BALB C
  • Oxidative Stress
  • Peroxiredoxins* / chemistry
  • Peroxiredoxins* / genetics
  • Peroxiredoxins* / immunology
  • Peroxiredoxins* / metabolism
  • Phylogeny
  • Protozoan Proteins* / genetics
  • Protozoan Proteins* / metabolism

Substances

  • Peroxiredoxins
  • Protozoan Proteins